Janovick Jo Ann, Knollman Paul E, Brothers Shaun P, Ayala-Yáñez Rodrigo, Aziz Abeer S, Conn P Michael
Division of Neuroscience, Oregon National Primate Research Center, Oregon Health and Science University, Beaverton, Oregon 97006, USA.
J Biol Chem. 2006 Mar 31;281(13):8417-25. doi: 10.1074/jbc.M510601200. Epub 2006 Jan 30.
Despite the prevalence of G protein-coupled receptors as transducers of signals from hormones, neurotransmitters, odorants, and light, little is known about mechanisms that regulate their plasma membrane expression (PME), although misfolded receptors are recognized and retained by a cellular quality control system (QCS). Convergent evolution of the gonadotropin-releasing hormone (GnRH) receptor (GnRHR) progressively decreases inositol phosphate production in response to agonist, validated as a measure of PME of receptor. A pharmacological chaperone that optimizes folding also increases PME of human, but not of rat or mouse, GnRHR because a higher percentage of human GnRHRs are misfolded structures due to their failure to form an apparent sulfhydryl bridge, and they are retained by the QCS. Bridge formation is increased by deleting (primate-specific) Lys191. In rat or mouse GnRHR that lacks Lys191, the bridge is non-essential and receptor is efficiently routed to the plasma membrane. Addition of Lys191 alone to the rat sequence did not diminish PME, indicating that other changes are required for its effects. A strategy, based on identification of amino acids that both 1) co-evolved with the Lys191 and 2) were thermodynamically unfavorable substitutions, identified motifs in multiple domains of the human receptor that control the destabilizing influence of Lys191 on a particular Cys bridge, resulting in diminished PME. The data show a novel and underappreciated means of posttranslational control of a G protein-coupled receptor by altering its interaction with the QCS and provide a biochemical explanation of the basis of disease-causing mutations of this receptor.
尽管G蛋白偶联受体作为激素、神经递质、气味分子和光信号的转导分子十分普遍，但关于调节其质膜表达（PME）的机制却知之甚少，尽管错误折叠的受体可被细胞质量控制系统（QCS）识别并保留。促性腺激素释放激素（GnRH）受体（GnRHR）的趋同进化导致其对激动剂的肌醇磷酸生成逐渐减少，这被确认为受体PME的一种衡量指标。一种优化折叠的药理学伴侣也能增加人GnRHR的PME，但对大鼠或小鼠的GnRHR则无此作用，因为较高比例的人GnRHR由于未能形成明显的巯基桥而呈错误折叠结构，并被QCS保留。删除（灵长类特异性的）赖氨酸191可增加桥的形成。在缺乏赖氨酸191的大鼠或小鼠GnRHR中，该桥并非必需，受体能有效转运至质膜。仅在大鼠序列中添加赖氨酸191并不会降低PME，这表明其发挥作用还需要其他变化。一种基于识别与赖氨酸191共同进化且在热力学上为不利取代的氨基酸的策略，在人受体的多个结构域中确定了基序，这些基序控制着赖氨酸191对特定半胱氨酸桥的不稳定影响，从而导致PME降低。这些数据显示了一种通过改变G蛋白偶联受体与QCS的相互作用来进行翻译后调控的新的且未被充分认识的方式，并为该受体致病突变的基础提供了生化解释。
